The document provides a comprehensive overview of microbial protein biosynthesis, detailing the structure, roles, and synthesis of proteins and the processes of transcription and translation. It explains the steps involved in gene expression, including initiation, elongation, and termination in transcription, as well as the phases of translation with associated components. Additionally, it compares protein synthesis in prokaryotes and eukaryotes, addressing regulation mechanisms of protein synthesis such as autoregulation and both positive and negative regulation.

1. MICROBIAL METABOLISM
Microbial Protein Biosynthesis

2. PROTEIN:
Proteins are large molecules composed of one or more
long chains of amino acids linked together by peptide
bonds and organized in a specific order .
The order is determined by the base sequence of
nucleotides in the genes that encode for proteins.
•Amino acids are the building blocks of protein.
•There are 20 different amino acids that combine to make
a protein.

3. Roles of Proteins:
•Structural component:
Forms scaffolding that maintains cell shape e.g. glycoprotein
•Transport:
Bacterial transport proteins mediate passive and active transport of
small solutes across membranes e.g. porins ( aquaporin),channels (Holin)
•Enzymes :
Enzymes are protein in nature and catalyze metabolic reactions e.g.
. RNA polymerase, ligase, helicase
•Motility:
Locomotory organs such as flagella and Pilli are also made of proteins
that is flagellin and pilin.
•Antibodies:
Antibodies are also protein in nature e.g. immunoglobulin IgG
•Proteins are also important in cell signaling , immune responses, cell adhe
and the cell cycle.

4. Central
dogma
RNA
Transcription
DNA
Proteins
Translation
Page 4

5. CENTRAL DOGMA

6. TRANSCRIPTION:
Transcription is the first step of gene expression, in which a particular
segment of DNA is copied into RNA (especially mRNA) by the enzyme RNA
polymerase.
If the gene transcribed encodes for a protein ,the result of transcription is
messenger RNA (mRNA), which Will then be used to create that protein via
the process of translation.
As in DNA replication ,DNA is read from 3` 5` during transcription.
Meanwhile ,complementary RNA is created from 5` 3` direction.
Only one of the two DNA strands ,called the template strand is used for
transcription.
The other DNA strand is called the coding strand ,because its sequence is
same as the newly synthesized RNA transcript (except for substitution of
Uracil for thymine).

7. Stages of Transcription:
Transcription is divided into
three stages
• Initiation
• Elongation
• Termination

8. INITIATION:
• In bacteria ,transcription begins with the binding of RNA
polymerase to the
promoter in DNA.
• Promoter sequences are DNA sequences that define where
transcription of
a gene by RNA polymerase begins.
• The promoter consists of two short sequences at -10 which is calle
pribnow box (TATAAT) and -35 positions (TTGACA) upstream
from the transcription start site.
• RNA polymerase is a core enzyme consisting of 5 subunits.
• At the start of initiation, the core enzyme is associated with a sigm
factor
that aids in the finding the appropriate -35 and -10base pairs
downstream
of promoter sequences.

9. INITIATION

10. ELONGATION :
• One strand of DNA , template strand (or non coding strand), is used as
template for RNA synthesis.
• As the transcription proceeds,RNA polymerase traverses the template stra
and uses Base pairing complementarity with the DNA template to create a
RNA copy.
• Although RNA polymerase traverses the template strand from 3` 5`h
coding (non template) strand and newly formed RNA can also be used as
reference points, So transcription can be described as occurring 5` 3
• This produces an RNA molecule from5` 3` an exact copy of the coding
strand (except that thymine are replaced with uracil ,and the nucleotides
are composed of ribose)
• Elongation also involves a proof reading mechanism that can replace
incorrectly incorporated Bases.

12. Termination:
Bacteria used two different strategies for termination:
Rho- independent
Rho- dependent
Rho- independent termination:
In this termination RNA transcription stops when the newly synthesized RNA
Molecule Forms a GC rich hairpin loop followed by a run of U's, which make
Detach from the DNA Template.
Rho-dependent termination:
In this type of termination, a protein factor called “Rho” destabilizes the
interaction between the template and mRNA , thus releasing the newly
synthesized mRNA from the elongation complex.

13. TERMINATION

14. Genetic code:
The genetic code is the set of all codons.
Codon:
An RNA base sequence corresponding to a particular amino acid is called
codon.
Properties of genetic codon:
The code is:
• universal
• Triplet
• Degenerative
• Non overlapping

15. TRANSLATION:
Translation involves decoding mRNA and covalently linking amino acids
together to form a polypeptide.
• This occurs at the ribosome.
Components Of Translation:
The translation system consists of four major components.
•Ribosomes
•tRNA
•Amino acyl tRNA synthetases
•Initiation , elongation and release factors
In prokaryotes all of these components are present throughout the cell.

16. Amino acid
attachment site

17. STAGES OF TRANSLATION:
Translation proceeds in four phases:
•Activation
•Initiation
•Elongation
•Termination

18. ACTIVATION:
•In activation the correct amino acid is covalently bonded to the correct tRN
•Activation is required for the translation to proceed.
•When the tRNA has an amino acid linked to it. It is called “charged”.
INITIATION:
•In prokaryotes initiation requires the large and small ribosome sub units,
the charged fMet-tRNA and three initiation factors and GTP.
•The 30s subunit attach to an mRNA molecule making use of the ribosome
binding site on the mRNA.
•The fMet- tRNA can then bind to the complex by base pairing of its anticod
with the AUG Codon on mRNA.
•This complex is called 30s initiation complex.
•The 50s subunit can now bind and the GTP is hydrolyzed in this energy
consuming step.
•This complex is called 70s initiation complex.

19. f- f-

20. ELONGATION:
Every addition of an amino acid to a growing polypeptide chain is the resu
an elongation cycle composed of three phases:
• Aminoacyl-tRNA Binding,
• The Transpeptidation Reaction
• Translocation
The process is aided by proteins called elongation factors (EF)
. The ribosome has three sites for binding tRNAs:
(1) the peptidyl or donor site (P site),
(2) the aminoacyl or acceptor site (A site), and
(3) the exit site (E site)
At the beginning of an elongation cycle, the P site is filled with fMet-tRNA
and the A and E sites are empty
AMINOACYL-tRNA BINDING:
The aminoacyl-tRNA corresponding to the codon in the A site is inserted
anticodon is aligned with the codon on the mRNA.

21. TRANSPEPTIDATION:
Transpeptidation is catalyzed by the Peptidyl Transferase. In this reaction,
amino group of the A site amino acid reacts with the carboxyl group of the
C-terminal amino acid on the P site tRNA. This results in the transfer of the
peptide chain from the tRNA in the P site to the tRNA in the A site, as a pep
bond is formed between the peptide chain and the incoming amino acid.
TRANSLOCATION:
Three things happen simultaneously:
(1) The ribosome moves one codon along mRNA so that a new codon is
(2) The peptidyl-tRNA moves from the A site to the P site
positioned in the A site
(3) The empty tRNA moves from the P site to the E site and subsequently lea
the ribosome

23. TERMINATION:
•Termination of the polypeptide happens when the A site of the ribosome
faces a stop codon(UAA,UGA,UGA).
•When this happens, no tRNA can recognize it but a releasing factor can
recognize nonsense codons and Causes the release of polypeptide chain
Events after termination:
•Hydrolyses of terminal peptidyl-tRNA bond
•Release of free polypeptide and the last uncharged tRNA from P site.
•The dissociation of 70s ribosome into 30s and 50s subunits.

24. TERMINATION

25. POLYRIBOSOMES:
•A single strand of mRNA is translated simultaneously by many ribosome
spaced Closely together.
•Such clusters of ribosomes are called polysomes or polyribosomes
•The use of polysomes is particularly advantageous to a cell in that the
Overall rate Of protein synthesis is increased compared to the rate that
would occur without Polysomes

26. Difference Between Protein
Synthesis In Prokaryotes And
Eukaryotes
PROKARYOTES
• mRNA is polycistronic
• mRNA processing is not
required(poly A tail and 7-
methyl GTP cap)
• coupled transcription –
translation
• The first amino acid is N-
formyl methionine.
• Ribosomes are smaller in
size (70S)
• Shine Delgarno sequence
is present
EUKARYOTES
• mRNA is monocistronic
• mRNA processing is
required(poly A tail and 7-
methyl GTP cap)
• No coupled transcription –
translation
• The first amino acid is
methionine.
• Ribosomes are larger in
size (80S).
• Shine Delgarno sequence
is not present

27. REGULATION OF PROTEIN SYNTHESIS
Protein synthesis regulation is of three types:
Autoregulation:
Protein itself inhibit initiation of transcription, and high concentration
of the protein will result in less transcription of mRNA that encodes the prote
Negative regulation:
In a negatively regulated system, an inhibitor is present in cell and prevent
transcription. This inhibitor is called repressor. An antagonist of the repres
an inducer is needed to allow initiation of transcription.e.g the E.coli lactose
system
Positive regulation:
In positive regulation a transcription factor is required to bind at the prom
in order to enable RNA polymerase to initiate transcription e.g. cAMP-CRP
Complex in the E.coli lactose system

28. AUTOREGULATION

29. POSITIVE AND NEGATIVE REGULATI

32. ANY QUESTION?